Journal of Membrane Science 279 (2006) 354–363

Electrospun polylactic acid nanofiber membranes assubstrates for biosensor assemblies

Dapeng Li a, Margaret W. Frey a,∗, Antje J. Baeumner b

a Department of Textiles and Apparel, Cornell University, Ithaca, NY 14850, United Statesb Department of Biological and Environmental Engineering, Cornell University, Ithaca, NY 14850, United States

Received 23 August 2005; received in revised form 12 December 2005; accepted 13 December 2005Available online 15 February 2006

Abstract

Biotin has been successfully incorporated into polylactic acid (PLA) nanofibers through electrospinning to prepare membrane substrates forbiosensors based on biotin–streptavidin specific binding. Biotin incorporated PLA nanofiber membranes were characterized with scanning electronmicroscopy (SEM), electron probe microanalysis (EPMA), and confocal microscopy. Under optimized conditions, small fiber size and uniformmorphology were achieved for PLA nanofibers with and without biotin incorporation. Sulfur mapping indicated a non-uniform distribution ofbem©

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iotin on the membranes, presumably due to aggregation of biotin during the electrospinning process. Pre-blocking the membranes effectivelyliminated non-specific binding between streptavidin and PLA. Preliminary biosensor assays confirmed that streptavidin immobilized on theembrane surface could capture a biotinylated DNA probe.2005 Elsevier B.V. All rights reserved.

eywords: Electrospinning; Nanofiber; Biotin; Streptavidin; Polylactic acid; Biosensor

. Introduction

A wide array of biosensors have been developed utilizing theapid, specific, and strong binding between biotin and strepta-idin [1]. In most cases, streptavidin is applied to a substrateaterial surface and subsequently coated with a biotinylated

iorecognition agent used to capture specific target analytes.he number of sites available for detection of the target analytes

s directly related to the surface area of the sensor substrate. Aariety of materials including gold surfaces of surface plasmonesonance (SPR) sensors [2–4], plastic films [5], and microflu-dic devices [6] have been used as substrates for sensors basedn biotin–streptavidin immobilization.

Researchers have recognized the advantage of increasing theurface area of the detector substrate to increase the number ofensing sites available without increasing the amount of overallample required [7,8]. Polymeric membranes with high surfacerea can be prepared by electrospinning. Electrospinning is aber formation process which relies on electrical rather than

mechanical forces to form fibers with sub micron diameters.These fibers (nanofibers) have exceptional properties due to theirminute diameter and large surface to mass ratio [9]. Non-wovenmats, collected via electrospinning have small pore size, highporosity, and large surface area. As a result, a small volumeelectrospun mat can provide a very large surface for sensing andeasy access for contaminants to the sensing sites. Although thepotential application of combining electrospun nanofiber mem-branes and biosensing has been recognized, limited studies havebeen done in this area [10,11].

Polylactic acid (PLA) has been successfully electrospunfrom a variety of solvents [12,13]. Additionally, a wide vari-ety of materials have been incorporated in electrospun PLAfibers to tailor the fibers for particular end uses. Nanoscaleclay particles have been incorporated in electrospun PLAfibers to control modulus and biodegradation rate for potentialbiodegradable packaging applications [14]. Carbon nanotubeshave been incorporated in electrospun PLA fibers for poten-tial use as bone graft materials [9]. Pharmaceutical chemicalshave been included in electrospun mats for controlled releasedelivery. Kenawy et al. [15] incorporated tetracycline hydrochlo-

∗ Corresponding author. Tel.: +1 607 255 1937/3151; fax: +1 607 255 1093.E-mail address: mfw24@cornell.edu (M.W. Frey).

ride in electrospun non-woven fabrics of poly(ethylene-co-vinylacetate) and PLA. Tetracycline hydrochloride incorporated

376-7388/$ – see front matter © 2005 Elsevier B.V. All rights reserved.oi:10.1016/j.memsci.2005.12.036

D. Li et al. / Journal of Membrane Science 279 (2006) 354–363 355

in the fibers was able to diffuse to the fiber surface overtime.

This paper details our efforts to incorporate biotin intoelectrospun PLA membranes by dispersing biotin in aPLA/chloroform/acetone solution prior to electrospinning. Elec-tron probe microanalysis (EPMA) confirms the presence ofbiotin in the electrospun fibers and that the final biotin levelsare proportional to the amount in the initial dispersions. Furtherexperimentation confirms that the biotin is fixed on the PLAfibers and cannot be washed off. Preliminary biosensor assaysfollowing the method described by Baeumner et al. [16] are usedto confirm that the nanofiber membrane can successfully immo-bilize streptavidin which in turn is used for the immobilization ofbiotinylated nucleic acid probes for the detection of a syntheticE. coli DNA.

2. Experimental

2.1. Materials

Polylactic acid (Mw = 186,000, Mw/Mn = 1.76) was sup-plied by Cargill Dow LLC (Minnetonka, MN). Chloroformand acetone were purchased from VWR Scientific (WestChester, PA). Both biotin and Streptavidin-Alexa 488 conju-gate (495 nm:519 nm excitation:emission) were purchased fromPierce Biotechnology Inc. (Rockford, IL). Phosphate bufferedsi

oCfbi(wNm

2

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a Branson 2510 ultrasonic cleaner (Branson Ultrasonics Corp.,CT) prior to adding PLA. Samples containing biotin were soni-cated for another 60 min immediately before electrospinning toinsure good dispersion of biotin. Detailed information on thesolutions and dispersions that were used in this study is shownin Table 1.

2.3. Electrospinning

The electrospinning apparatus consisted of a programmablesyringe pump (Harvard Apparatus, MA) and a high-voltage sup-ply (Gamma High Voltage Research Inc., FL). Variations of theelectrospinning conditions are also summarized in Table 1. Alu-minum foil was used as the collector in all cases except for theelectrospinning of the membranes for biosensor assay.

2.4. Instrumental analyses

Morphology and fiber size of electrospun PLA nanofiberswere examined with a Leica 440 scanning electron microscope(SEM) after being coated with Au–Pd or with a LEO 1550 fieldemission scanning electron microscope (FESEM).

Biotin distribution on nanofiber membranes was character-ized with a JEOL 8900 electron probe microanalyzer (EPMA).Bllbrii

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aline (PBS) and Tween 20 were purchased from Aldrich Chem-cal Co. (Milwaukee, WI).

All general chemicals and buffer reagents (reagent grader above) for biosensor assay were purchased from Sigmaompany (St. Louis, MO). Organic solvents were purchased

rom Aldrich Chemical Co. (Milwaukee, WI). Predator mem-ranes were obtained from Pall/Gelman Company (Port Wash-ngton, NY). Lipids were purchased from Avanti Polar LipidsAlabaster, AL). Sulforhodamine B (SRB) and streptavidinere acquired from Molecular Probes Company (Eugene, OR).ucleic acid probes and synthetic targets, with their respectiveodifications, were purchased from Qiagen (Valencia, CA).

.2. Preparation of electrospinning solutions/dispersions

PLA and in chloroform/acetone solvent (3:1 volume ratio)ere mixed over night on an InnovaTM 2300 platform shaker

New Brunswick Scientific Co., NJ). For samples containingiotin, biotin was dispersed in chloroform/acetone solvent using

able 1omposition of electrospinning solutions/dispersions and electrospinning cond

un PLA (wt%) Biotin (wt%, relative to PLA) Voltage (k

-0-1 6 0 15-0-2 6 0 15-0-3 6 0 15-0-1 8 0 15-0-2 8 0 15-1 8 1 15-2 8 2 15-3 8 3 15

oth energy dispersive X-ray spectroscopy (EDS) and wave-ength dispersive X-ray spectroscopy (WDS) were used to col-ect characteristic K� X-ray emission of sulfur atoms in theiotin molecules. As a semi-quantitative means, Ratemeter X-ay counting was used to correlate the amount of biotin in thenitial dispersion with the amount that has been incorporatednto the membranes.

A Leica TCS SP2 laser confocal scanning microscope wassed to track the biotin–streptavidin specific binding by imag-ng the fluorescence of streptavidin-Alexa 488 conjugate treatedanofiber membranes. Scanning was performed starting fromhe top layer of each membrane towards the deeper layers andtopped when no fluorescence could be detected. To preparehe samples for confocal analysis, 50 �L of diluted streptavidin-lexa 488 conjugate solution was applied to a ∼4 mm × 4 mmanofiber membrane that was pre-wet out with PBS/Tween 20uffer. Treated membranes were subjected to repeated wash withBS/Tween 20 buffer to remove any extra streptavidin-Alexa88 conjugate before being placed onto a glass slide with a coverlass for confocal analysis.

Feed rate (�L/min) Needle size (mm) Ground distance (cm)

10 0.26 105 0.26 10

10 0.41 1010 0.41 12

5 0.41 1210 0.41 1210 0.41 1210 0.41 12

356 D. Li et al. / Journal of Membrane Science 279 (2006) 354–363

An Imass CAA2 Contact Angle Analyzer (Accord, MA) wasused to measure the contact angle between deionized H2O andboth electrospun PLA and PLA-biotin membranes.

SEM image analysis was conducted using Scion Image Beta4.02 (Scion Corporation, Frederick, MD, www.scioncorp.com)to obtain the size range of nanofibers for each membrane.

2.5. Biosensor assay

For biosensor assay, PLA membranes were electrospun onto acopper-backed laminate from a solution of 8 wt% PLA in the sol-vent chloroform/acetone (3:1 volume ratio) and an applied volt-age of 15 kV. The PLA membranes were cut into 3 cm × 0.4 cmstrips and pre-wet with PBS/Tween 20 buffer. Twenty picomolesstreptavidin dissolved in Na2CO3/NaHCO3 buffer, pH 9.0, con-taining 5% methanol (20 pmol/�L) was deposited on PLA stripsto form a capture zone. The membranes were put into the vac-uum oven at 103.421355 × 103 Pa (15 psi) and 50 ◦C for 1 h.The entire membrane was then blocked with a blocking reagent(0.02% polyvinylpyrollidone, 0.5% gelatin, and 0.005% caseinin Tris Buffered Saline) to prevent any possible non-specificbinding between other proteins and the membrane and dried ina vacuum oven at 36 ◦C and 103.421355 × 103 Pa (15 psi) for2.5 h. Such membrane strips were then assembled with laminateaccording to the design shown in Scheme 1. The capture zonewftss

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was inserted into a test tube containing 10 �L of either positiveor negative sample. After the sample was allowed to wick up themembrane strip, 35 �L of washing buffer (40% formamide, 8×SSC, 0.2% Ficoll, 0.2 M sucrose) were added to the test tubesto provide enough liquid to wet out the entire length of the strip.The color of the strip was measured with a BR-10 reflectometer(ESECO Company, Cushing, OK) to detect the presence of SRBin the capture and background zones.

3. Results and discussion

3.1. Electrospun PLA nanofibers

SEM images of PLA nanofibers electrospun under differentconditions are shown in Fig. 1. Sample 6-0-1 (Fig. 1a) shows atypical beaded-fiber morphology, which is mostly attributed tothe relatively low concentration of PLA in the electrospinningsolution. The beaded-fiber morphology is not affected by feedrate even when the feed rate is decreased by a factor of two,as seen from the image of sample 6-0-2 (Fig. 1b). The fiberuniformity improves slightly when the nozzle size is increasedwith other conditions held the same. Nanofibers electrospun witha needle of 0.41 mm in diameter are shown in Fig. 1c. Underthese conditions, the bead-fiber morphology has been replacedby spindle-fiber morphology.

A noticeable change in uniformity occurs when the concen-tdctf(tcwtbsia

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as 2 cm away from the free end. For comparison, polyethersul-one (PES) membranes were also used in the biosensor assay. Inhis case, the entire strip is a 7 cm long PES strip and 20 pmoltreptavidin was deposited 2 cm away from one end using theame procedure as for the PLA membranes.

The biosensor assay was conducted according to the protocoleveloped by Baeumner et al. [16]. In brief, biotinylated capturerobes (5′-CCg TTg gCA CAg CAA ATA-3′), universal reporterrobes (5′-gTC Tgg TgA ATT ggT TCC ggg ggg Tgg ggg Tgggg Tgg-3′) and SRB-entrapping liposomes bearing a universalrobe (5′-CCA CCC CCA CCC CCA CCC CC-3′) at their outerembrane were mixed in a hybridization buffer. A synthetic

arget DNA (5′-ggC AAC CgT gTC gTT TAT CAg ACC ACTAA CCA Agg C-3′) derived from the E. coli clpB mRNA wasdded to the mixture (H2O in the case of negative controls)nd incubated at 41 ◦C for 20 min. The exact composition ofhe mixture was: 2 �L liposomes, 1 �L reporter probe (2 pmol),�L target sequence (500 fmol), 1 �L capture probe (1 pmol),nd 5 �L hybridization buffer (20% formamide, 4× SSC, 0.4%icoll type 400, 0.4 M sucrose). One end of a membrane strip

Scheme 1. Design of the strip for biosensor assay.

ration of PLA is increased from 6% to 8% and the groundistance is increased by 2 cm, as seen in Fig. 1d, without signifi-antly increasing fiber size. Fiber diameters range from 200 nmo 5 �m. The fiber size can be made slightly smaller when theeed rate is decreased from 10 to 5 �L/min at 8% concentrationcompare Fig. 1d with Fig. 1e), but decreasing feed rate doubleshe time for electrospinning the same amount of nanofibers. Thehange in fiber size as a function of PLA concentration agreesith previous studies [17]. Fibers reported here were spun from

he same solvent system with the same polymer concentrationut using a larger molecular weight PLA (Mw = 186,000 ver-us Mv = 48,000). Increased molecular weight is associated withncreased viscosity of the spinning solution, which in turn isssociated with increased electrospun fiber diameter [18].

Fig. 2 shows the SEM images of PLA nanofibers spun fromolutions with different amounts of biotin dispersed in the spin-ing solution. Fiber diameters ranged from 150 nm to 5 �m aseasured by image analysis and were independent of biotin

oncentration. The addition of biotin in the amount of up to% relative to PLA weight in the electrospinning dope does notake a significant difference in fiber size and uniformity.

.2. EPMA analysis

EDS spectra of PLA nanofiber membranes with and with-ut biotin are shown in Fig. 3 before and after being subjectedo PBS buffer wash. Enrichment of sulfur is observed in PLA-iotin nanofibers. Because sulfur atoms are solely attributed toiotin molecules in this system (see Scheme 2 for the structure ofiotin molecules), the presence of the sulfur peak in EDS spec-ra indicates the successful incorporation of biotin molecules in

D. Li et al. / Journal of Membrane Science 279 (2006) 354–363 357

Fig. 1. SEM images of electrospun PLA nanofibers. (a) 6-0-1, (b) 6-0-2, (c) 6-0-3, (d) 8-0-1, and (e) 8-0-2.

nanofibers through electrospinning. Secondly, the sulfur peakshown in Fig. 3d indicates that biotin is not removed when themembrane is subjected to PBS buffer wash. Also seen from Fig. 3is that buffer wash causes two major changes in the spectra: theincreased background and the appearance of some new peakssuch as Na, Cl, K, and P. These peaks are introduced by PBS

buffer wash and do not overlap with the sulfur peak located at2.3 keV.

Further characterization using X-ray counting of PLAnanofiber membranes with and without biotin is shown in Fig. 4.The X-ray counts of the sample without biotin are the back-ground signal level [19]. Subtracting the background, the X-ray

358 D. Li et al. / Journal of Membrane Science 279 (2006) 354–363

Fig. 2. SEM images of biotin incorporated electrospun PLA. (a) 8-1, (b) 8-2, and (c) 8-3 (refer to Table 1 for sample information).

counts of the biotin incorporated PLA samples are proportionalto the amount of biotin dispersed in the electrospinning dopes,which provides us a semi-quantitative estimate of the level ofbiotin incorporated into the membranes. Comparing the X-raycount of the membrane before PBS buffer wash and that after, wecan see a much higher background generated in the membraneafter PBS wash (260 versus 667). Subtracting the backgroundfrom the control that is subjected to PBS wash, the PLA mem-

brane electrospun from the dispersion with 3% biotin shows thatabout 2/3 of the amount of biotin initially dispersed in the spin-ning dope remains on the membrane. The reason for examiningthe biotin retention on the membrane after PBS wash is to con-firm that biotin will not be removed by PBS buffer wash duringbiosensor assays.

To examine the distribution of biotin on nanofiber mem-branes, sulfur mapping was performed using EPMA. In Fig. 5,

Fwf

ig. 3. EDS spectra of electrospun PLA nanofiber membranes (a) 8-0-1 and (b) 8-3ash (see Ref. [16]). Spectrum (b) was obtained by peeling the membrane off the al

oil on the back of the membranes.

are before PBS buffer wash, while (c) 8-0-1 and (d) 8-3 are after PBS bufferuminum foil collector while the rest spectra were obtained with the aluminum

D. Li et al. / Journal of Membrane Science 279 (2006) 354–363 359

Fig. 3. (Continued ).

the increased number of blue spots in the PLA-biotin nanofibermembrane over the PLA only membrane confirms dispersion ofbiotin throughout the PLA-biotin nanofiber membrane. On theother hand, several large bright spots on the EPMA map of thePLA-biotin nanofiber membrane indicate that biotin is hetero-geneously distributed on the mapping area. EMPA mapping of

Fig. 4. Sulfur X-ray counts as a function of biotin concentration in electro-spinning dopes. At least eight areas (each area about 500 �m × 700 �m) arerandomly selected for sulfur K� X-ray counting for each nanofiber membranesample. Sixty seconds is applied as counting time under a current of 56 nA.

sulfur content in one of the bright spots is shown in Fig. 6 at ahigher magnification. Under this magnification it is evident thatthe distribution of sulfur signals matches the fibers alignment.This pattern match further confirms the incorporation of biotininto PLA nanofibers.

3.3. Confocal microscopy analysis

To confirm that biotin incorporated in PLA nanofiber mem-branes is available to bind with streptavidin, specific binding

Scheme 2. Structure of biotin.

360 D. Li et al. / Journal of Membrane Science 279 (2006) 354–363

Fig. 5. Comparison of the sulfur map of PLA nanofiber membranes with (top, 8-3) and without (bottom, 8-0-1) biotin. Refer to Table 1 for sample information.

between the incorporated biotin and a fluorescent dye-Alexa488-labeled streptavidin conjugate was studied via confocalmicroscopy. Fig. 7 shows the top view of the overlapped flu-orescence images of both a PLA nanofiber membrane and thePLA-biotin nanofiber membrane. Also shown in Fig. 7 is thefirst frame of the optical images of both membranes. The colorbar beside each image indicates the location of fluorescencesignals on the scanned layers. The purple and blue nanofibersare located on the layers closest to the surface of the mem-brane, while the red and pink nanofibers are located on thelayers farthest away from the surface. The PLA-biotin mem-brane exhibits greater fluorescence at all depths than the PLAonly membrane. More importantly, however, fluorescence on thePLA-biotin membranes follows the pattern of the electrospunfibers. Biotin incorporated in the fibers appears to be success-fully bound to fluorescence labeled streptavidin along the lengthof the fibers.

The difference between PLA membrane and PLA-biotinmembrane is also seen in the profile images shown in Fig. 8band c. The travel distance of fluorescence is about twice as longin the biotin incorporated PLA membrane than in the pure PLAmembrane. On the other hand, the control membrane treatedonly with PBS buffer (Fig. 8a) shows no fluorescence at all.

Such difference is certainly due to the incorporation of biotin.First of all, the florescence resulted from the pure PLA mem-brane is simply caused by the non-specific binding betweenstreptavidin and the substrate, or the physical absorption of theprotein onto the membrane (Fig. 8b). The incorporation of biotininto PLA membrane on the one hand introduces the active sitesfor specific binding with streptavidin while on the other handdecreases the hydrophobicity of the PLA membrane (see thecontact angle measurements in Fig. 9). Both the introductionof active binding sites and the increased hydrophilicity of themembrane contribute to the increase in streptavidin penetra-tion.

Also shown in Fig. 8 is the difference in fluorescence betweenblocked and non-blocked PLA membranes. Comparison ofFig. 8b and d confirms that pre-blocking have effectively sup-pressed the non-specific binding of streptavidin-Alexa 488 con-jugate. As seen from Fig. 8d, blocked PLA membrane has littleor no fluorescence, the same appearance as the control (Fig. 8a)sample. Blocked PLA-biotin membrane (Fig. 8e), on the otherhand, has fluorescence, indicating that biotin is accessible to bindwith streptavidin after the membrane is blocked. Blocked PLA-biotin membrane shows approximately the same penetrationdepth along the profile as does the sample without pre-blocking

D. Li et al. / Journal of Membrane Science 279 (2006) 354–363 361

Fig. 6. High magnification sulfur mapping of biotin incorporated PLA nanofiber membrane.

(Fig. 8c). Fig. 8d and e clearly shows the difference betweenPLA and PLA-biotin membranes when non-specific binding iseffectively suppressed by pre-blocking.

The confocal results indicate two important properties ofthe PLA-biotin nanofiber membranes. First, fluorescence sig-nal appears along the length of individual fibers, indicatingthat biotin incorporated into individual fibers is accessible forstreptavidin binding. The heterogeneous distribution of biotinevident from EPMA mapping (Fig. 5) does limit the availabil-ity of sites for biotin–streptavidin binding. Second, althoughthe biotin incorporated into PLA membrane is accessible tostreptavidin to bind with, only the biotin located approximately10 �m depth of the entire thickness of the membrane hasspecifically bound with streptavidin in confocal experiments.The hydrophobicity of PLA nanofibers may limit the wick-ing of streptavidin through the membrane under aqueous bufferenvironment.

3.4. Biosensor assay

To demonstrate that the analyte solution can travel throughnanofiber membranes and that the target E. coli DNA can be

captured by streptavidin on the nanofiber surface a biosensorassay was performed. An electrospun PLA nanofiber mem-brane was used as the substrate as shown in Scheme 1. Fil-ter paper is attached to one end of the strip to collect excessliposomes that are not specifically bound with the depositedstreptavidin during the wicking process. For comparison, aparallel biosensor assay using regular PES membrane as thesubstrate was also performed. Shown in Fig. 10 are the pho-tographs of typical biosensor assay results from both membranesubstrates. The test solution wicked successfully through thePLA nanofiber membrane, capture probes were successfullytrapped where streptavidin had been deposited and excess solu-tion wicked past the capture zone to the filter paper. Specificsignals for the clpB synthetic target sequence in the capturezones were 57 for the PES sensor and 69 for the PLA sen-sor. Signals of the corresponding background zones were 26and 59. This indicates that streptavidin was successfully immo-bilized on the PLA membrane through deposition and thatit was capable of capturing biotinylated DNA capture probeshybridized to the target sequence-liposome complex, thus prov-ing the validity of our concept in nanofiber membrane-basedbiosensors.

362 D. Li et al. / Journal of Membrane Science 279 (2006) 354–363

Fig. 7. Pattern match between the distribution of fluorescence and that of PLA nanofiber membranes with (bottom, 8-3) and without (top, 8-0-1) biotin. Refer toTable 1 for sample information.

Fig. 8. Fluorescence image profiling of streptavidin-Alexa 488 conjugate treated PLA and PLA-biotin nanofiber membranes. (a) Non-blocked PLA membrane appliedonly with PBS buffer (control). (b) Non-blocked PLA membrane treated with streptavidin-Alexa 488 conjugate. (c) Non-blocked PLA-biotin (8–3%) membranetreated with streptavidin-Alexa 488 conjugate. (d) Blocked PLA membrane treated with streptavidin-Alexa 488 conjugate. (e) Blocked PLA-biotin (8–3%) membranestreated with streptavidin-Alexa 488 conjugate.

D. Li et al. / Journal of Membrane Science 279 (2006) 354–363 363

Fig. 9. Contact angle differences of the electrospun PLA membranes with andwithout biotin.

Fig. 10. Capture of E. coli synthetic target sequence on the PES membrane (top)and electrospun PLA nanofiber membrane (bottom). Circled area is the capturezone where streptavidin is deposited. Squared area is where the backgroundmeasurement is taken that serves as an internal control of the performance ofeach membrane, i.e. presenting possible non-specific binding of liposomes onthe membrane.

4. Conclusions

Biotin has been successfully incorporated into PLAnanofibers through electrospinning without significantly chang-ing the morphology and size of the resulting nanofibers.Nanofiber membranes were characterized with EPMA to con-firm biotin incorporation in the fibers. Although biotin is dis-persed throughout the nanofiber membranes the distribution isheterogeneous and regions of locally high biotin concentrationare evident. Confocal microscopy analysis indicates that bothspecific and non-specific binding occur between streptavidinand the biotin incorporated PLA nanofiber membranes. Withblocking reagent, non-specific binding can be suppressed effec-tively. Fluorescence labeled streptavidin binds along the surfaceof PLA-biotin fibers providing evidence that biotin incorporatedvia electrospinning is available at the fiber surface. Biosen-sor assay experiments confirm that the PLA nanofiber mem-branes can successfully transport analyte solutions via wick-ing. A biotinylated DNA probe was successfully captured byimmobilized streptavidin in a preliminary biosensor assay usingan electrospun PLA nanofiber membrane as substrate. Furtherefforts are being made in (1) optimizing the nanofiber substrateby decreasing the fiber diameter and increasing biotin loadingand (2) functionalizing the electrospun nanofibers for specificcapture and detection of environmental pathogens or other con-taminates.

Acknowledgements

The project was supported by the National Research Initiativeof the USDA Cooperative State Research, Education and Exten-sion Service (grant number 2005-35603-15298). The authorsacknowledge the contribution made by Ms. Barbara Leonardand Ms. Jamie Mullally.

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